Field of the Invention
The present invention is related to implanted compositions with vascular endothelial growth factor (VEGF) in combination with other growth factors.
Background of the Invention
A healthy blood circulatory system is pivotal to the development and maintenance of functional tissues and organs; damaged blood vessels or compromised circulation can result in ischemic tissue with limited intrinsic regeneration potential (Kraehenbuehl et al., Biomaterials vol. 30, no. 26, pp. 4318-24, 2009). The ideal treatment for vascular damage is the regeneration of autologous blood vessels, which can be optimized by cell engineering (Srivastava et al. Nature vol. 441, no. 7079, pp. 1097-9, 2006; Adams et al., Trends in Cardiovascular Medicine, vol. 17, no. 7, pp. 246-51, 2007; Sales et al., Trends Biotechnol, vol. 23, no. 9, pp. 461-7, 2005) and extracellular matrix manipulation (Sun et al., Regenerative Medicine, vol. 3, no. 3, 435-47, 2009; Yang et al., Tissue Engineering vol. 7, no. 6, 679-89, 2001; Ravi et al., Regenerative Medicine vol. 5, no. 1, 107-20, 2010). Vascular regeneration, however, remains a complex process which includes the mobilization, chemotaxis, adhesion, proliferation, and differentiation of progenitor cells (Guiducci et al., vol. 47, suppl. 5, pp. v18-20, 2008); further, the process is influenced by tissue composition and, possibly, inflammation induced by tissue injury.
Research has uncovered a broad range of pharmaceutical and biomedical applications for polymeric hydrogels, due to their three-dimensional (3D) structural and mechanical similarity to the native ECM of many tissues. Optimally tailored biodegradable polymer hydrogel scaffolds could provide a constant delivery of bioactive angiogenic factors, advancing therapeutic vascularization. Moreover, these porous hydrogel scaffolds permit circulating cells to infiltrate into them to degrade them, thereby releasing entrapped growth factors (GFs) and facilitating neovascularization (Sheridan et al., J Controlled Release, vol. 64, no. 1-3, pp. 91-102, 2000; Perets et al., J Biomed Mater Res Part A, vol. 65A, no. 4, pp. 489-97, 2003). For in vivo transplantation purposes, the hydrogel scaffold should have enough mechanical strength to maintain the integrity of its porous structure during the vascularization process. Overall, a porous hydrogel scaffold capable of a constant release of GFs, proper mechanical strength, and rapid degradation may enable vascular engineering.
Polysaccharide-based hydrogels are non-toxic and biodegradable. From a structural point of view, polysaccharides have reactive functional groups that can be modified to form hydrogels with specific characteristics of interest. In recent years, hyaluronan-based hydrogels have been extensively investigated as scaffolds to stimulate in vivo angiogenesis (Riley et al., Biomaterials vol. 27, no. 35, pp. 5935-43, 2006; Elia et al., Biomaterials, vol. 31, no. 17, pp. 4630-8, 2010; Hosack et al., Biomaterials vol. 29, no. 15, 2336-47, 2008; Pike et al., Biomaterials, vol. 27, no. 30, pp. 5242-51, 2006); these hydrogels were manipulated to deliver different angiogenic factors to improve neovascularization. Lee et al. demonstrated that a VEGF-encapsulated alginate hydrogel scaffold promoted angiogenesis (Lee et al., Nature, vol. 408, no. 6815, 998-1000, 2000). However, they reported that little to no tissue or blood vessel ingrowth into these hydrogel scaffolds occurred. Recently, the Cohen group reported that the in vivo prevascularization of an alginate scaffold improved therapeutic vascularization (Dvir et al., Proc Natl Acad Sci USA, vol. 106, no. 35, 14990-5, 2009). Although dextran-based hydrogels have been investigated for various purposes (Bos et al., Biomaterials vol. 26, no. 18, pp. 3901-9, 2005; Ferreira et al., Biomaterials vol. 28, no. 17, pp. 2706-17, 2007), their potential for in vivo therapeutic vascularization has not been fully explored.
Although many different types of polymeric hydrogels have been developed since the 1950s (Kopecek, J. Nature 2002, vol. 417, pp. 388-391), they all fall into one of two basic categories of polymer: natural or synthetic. Natural polymers have gained interest over the past few decades because of their biocompatibility and the presence of biologically recognizable groups to support cellular activities (Van Tomme et al. Expert Rev. Med. Dev. 2007, vol. 4, pp. 147-164). Among the natural polymers, dextran is a colloidal, hydrophilic, biocompatible, and nontoxic polysaccharide composed of linear α-1,6-linked D-glucopyranose residues with a low fraction of α-1,2, α-1,3 and α-1,4 linked side chains. Also, dextran can be biodegraded by dextranase, which exists in mammalian (including human) tissues. From a structural point of view, dextran has reactive hydroxyl groups (i.e. —OH) that can be modified to form hydrogels via crosslinking by photochemical and other means. As dextran is naturally resistant to protein adsorption and cell adhesion, modification of its polymer backbone allows development of a hydrogel with specific characteristics. Because of these properties, dextran and its hybrids have been extensively investigated as drug and/or gene carriers. For examples, dextran-based biomaterials have been employed in cell immobilization (Massia et al., Biomaterials, 2000, vol. 21, pp. 2253) and gene transfection (Azzam et al., Macromol. Symp., 2003, vol. 195, p. 247) and as carriers for a variety of pharmaceutically active drugs (de Jong et al., Macromolecules, 2000, vol. 33, p. 3680; Kim et al., J. Biomater. Appl., 2000, vol. 15, p. 23; Won et al., Carbohydr. Polym., 1998, vol. 36, p. 327; Kim et al., Arch. Pharma. Res., 2001, vol. 24, p. 69; Chu, C. C., in: Biomaterials Handbook—Advanced Applications of Basic Sciences, and Bioengineering, D. L. Wise (Ed.), p. 871. Marcel Dekker, New York, N.Y. (2003); Won et al., in: Biomaterials & Engineering Handbook, D. L. Wise (Ed.), p. 356. Marcel Dekker, New York, N.Y. (2000); Zhang et al., J. Biomater. Appl., 2002, vol. 16, p. 305; Peppas et al., Europ. J. Pharma. Biopharma., 2000, vol. 50, p. 27; Van Tomme et al., Biomaterials, 2006, vol. 27, p. 4141).
Many attempts have been made to engineer dextran-based polymers for various applications (Heinze et al., In Polysaccharides Ii, Springer-Verlag Berlin: Berlin, 2006; p. 199). Van Tomme et al. recently reviewed both chemically and physically crosslinked dextran-based hydrogels that were developed for protein release (Van Tomme et al. Expert Rev. Med. Dev. 2007, vol. 4, pp. 147-164). To generate chemically crosslinked dextran hydrogels, the major modification challenge is to introduce polymerizable bonds for efficient crosslinking. A common approach is to incorporate vinyl groups via different types of acrylates, thus enabling photocrosslinking. Such acrylates include glycidyl acrylate (Edman, et al., I. J. Pharm. Sci. 1980, vol. 69, pp. 838-842), glycidyl methacrylate (Vandijkwolthuis et al., Macromolecules, 1995, vol. 28, pp. 6317-6322), methacrylate (Kim et al., J. Biomed. Mater. Res., 2000, vol. 53, pp. 258-266; Ferreira et al., Biomaterials, 2007, vol. 28, pp. 2706-2717), acrylate (Zhang et al., J. Polym. Sci. Polym. Chem., 1999, vol. 37, pp. 4554-4569) and hydroxyethyl methacrylate (vanDijkWolthuis et al., Macromolecules, 1997, vol. 30, pp. 4639-4645; vanDijkWolthuis et al., Polymer, 1997, vol. 38, pp. 6235-6242). These hydrogels were proven to be efficient protein carriers. Chu et al. also developed maleic-anhydride- and allyl-isocyanate- (AI-) based dextran hydrogels (Kim et al., J. Biomed. Mater. Res., 2000, vol. 53, pp. 258-266; Zhang et al., J. Polym. Sci. Polym. Chem., 2000, vol. 38, pp. 2392-2404), which were shown to have tunable properties. Other than UV photocrosslinking, the Schiff reaction has also been employed to form crosslinks by oxidizing dextran rings into aldehyde groups (Maia et al., “Synthesis and characterization of new injectable and degradable dextran-based hydrogels,” Polymer, 2005, vol. 46, pp. 9604-9614; Ito et al., Biomaterials, 2007, vol. 28, pp. 3418-3426).
One approach to preparing dextran-based hydrogels involves the use of a synthetic polymer precursor so that the resulting hydrogels can have both synthetic and naturally occurring polymers within a single entity. Among synthetic polymer precursors that couple with dextran, poly(ethylene glycol) (PEG) is popular because it is a unique amphiphilic, biocompatible but non-biodegradable polymer, and has been explored for many biomedical applications. Although PEG is not biodegradable, lower molecular weight PEG can be readily excreted from the body via kidney and liver, thereby making it more suitable for drug delivery. In addition, PEG has also been employed to improve biocompatibility (Zhang et al., Biomaterials, 2002, vol. 23, p. 2641-2648; Chung et al., Int. J. Biol. Macromol., 2003, vol. 32, p. 17), promote peptide immobilization (Hem et al., J. Biomed. Mater. Res., 1998, vol. 39. p. 266; Wang et al., J. Membr. Sci., 2002, vol. 195, p. 103), prolong protein drug circulating time (Koumenis et al., Int. J. Pharma., 2000, vol. 198, p. 83; Greenwald et al., Adv. Drug Deli. Rev., 2003, vol. 55, p. 217), increase bioactivity (Muslim et al., Carbohydr. Polym., 2001, vol. 46. p. 323-330) and reduce immunogenicity (Hu et al., Int. J. Biochem. Cell. Biol., 2002, vol. 34, p. 396-402).